The ventilatory responses to respiratory and metabolic acid-base disturbances are primarily mediated through stimulation of the central chemoreceptors which are thought to be located close to the surface of the ventrolateral medulla. Characterization of central chemosensitivity has heretofore been obtained primarily from intact preparations which do not allow distinction to be made between primary chemosensitive cells and those which are secondarily excited. The general objective of this proposal is to define the stimulus-response characteristics of individual chemosensitive neurons by using tissue culture techniques. This approach has been successfully utilized for the characterization of thermosensitive neurons in the hypothalamus. Preliminary studies using these techniques on ventral medullary tissue have shown that acid-sensitive neurons are readily identifiable in tissue culture. Three hypotheses regarding the nature of chemosensitivity in tissue culture are proposed for study. (1) It is proposed that acid sensitivity in tissue culture is specific to ventral medulla. This will be tested by comparing the stimulus-response characteristics of spontaneously active neurons in ventral and dorsal medullary tissue cultures. (2) It is proposed that a portion of the ventral medullary neurons are intrinsically ("primarily") acid sensitive. This will be tested by determining whether chemosensitivity persists in some neurons after blockade of synaptic transmission (high Mg++-low Ca++ perfusate). (3) It is proposed that acetylcholine and opiates modulate the acid sensitivity of chemosensitive neurons. This will be tested by determining the effect of agonists and antagonists of ACh and opiates on the stimulus response of acid-sensitive neurons. Studies will be performed on 21-35 day tissue cultures prepared from ventral and dorsal medullary tissue of newborn rats. Extracellular recordings will be made while the perfusate pH is randomly varied by either altering PCO2 at constant HCO3- or altering (HCO3-) at constant PCO2. Automated data collection will allow computation of linear regression of neuronal firing rate and ECF pH.